RESPONSE OF PHOSPHATIDYLCHOLINE IN THE GEL AND LIQUID-CRYSTALLINE STATES TO MEMBRANE-SURFACE CHARGES

The influence of membrane surface charge on the conformation of the choline head group of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) was investigated in the gel and liquid-crystalline states by using H-2 NMR spectroscopy of specifically choline-deuterated DMPC. The surface charge was made progressively more negative through admixture of various proportions of 1,2-dimyristoyl-sn-glycero-3-phosphoglycerol (DMPG). All membrane compositions showed nearly identical gel- to liquid-crystalline-phase transitions centered about 24-degrees-C. The gel-state H-2 NMR spectra from all three choline head-group deutero-labeling positions (alpha, beta, and gamma) decreased in intensity and broadened relative to the liquid-crystalline-state spectra. These effects were not so severe that they masked the overriding influence of surface charge on the choline head-group conformation as reflected in the H-2 NMR spectra. Thus, in both the liquid-crystalline and gel states, the presence of negative surface charge caused the quadrupole splitting from DMPC-alpha-d2 to increase while causing that from DMPC-beta-d2 and DMPC-gamma-d9 to decrease. These effects were progressive with increasing density of negative surface charge. Correlation plots of the quadrupole splittings obtained, under otherwise identical conditions, from different deutero-labeling positions were linear over most of the range of surface charge densities, in both the liquid-crystalline and gel states, for all three correlations (alpha-beta, beta-gamma, and alpha-gamma). At extreme surface charge densities, the alpha-beta and alpha-gamma correlations showed biphasic behavior in that, at high surface charge densities, the change in the quadrupole splittings from DMPC-alpha-d2 became less pronounced. The beta-gamma correlation was linear over the entire range of surface charge densities, regardless of the phase state of the membrane. In terms of the present model of the conformational change undergone by the choline group in response to membrane surface charges, the biphasic alpha-beta and alpha-gamma correlations for negative surface charges indicate a steric hindrance to the tilting of the choline P-N vector toward the membrane surface. These results demonstrate the utility of the H-2 NMR approach for the investigation of surface charge effects in gel-state lipid membranes and indicate the potential for its exploitation in the study of other membrane-associated phenomena such as lipid lateral-phase segregation. In addition, we describe a model of the relationship between surface charge and H-2 NMR quadrupole splittings which reproduces both qualitatively and quantitatively all the major features as established experimentally. On the basis of the premise that the choline group tilts in response to surface charges, the model provides a context for understanding the particulars of the H-2 NMR response, and provides new predictions subject to experimental verification.